Our principal aim is to examine the role of regulatory mutations in determining the anatomical and physiological variations among individuals, populations, and, particularly, species of vertebrates. It is proposed that lysozyme provides an excellent model system to study the molecular basis for adaptive change at the organismal level. Our discovery that the stomach mucosa of ruminants contains an extraordinarily high concentration of lysozyme provides insight into the origin of the ruminant mode of digestion. It appears from our preliminary work that both structural and regulatory mutation affecting the lysozyme gene have been responsible in part for this major evolutionary change in digestive physiology and life style. We plan, therefore, to make a thorough comparison of the lysozymes and the lysozyme genes of both ruminant and non-ruminant mammals. A parallel study will be made with birds, for which there is also evidence of enormous species differences in the amount of lysozyme produced by a given cell type. The main techniques to be used are those traditionally used in this laboratory for protein comparisons (namely, enzymological, immunological, and amino acid sequencing techniques) as well as in vitro protein synthesis and the newer techniques for nucleic acid comparison, including annealing, restriction mapping, and sequencing of lysozyme-specific DNA obtained by cloning of both cDNA an genomic DNA. Similar techniques will be used to analyze the polymorphism found in duck lysozymes. The multiple lysozymes appear to be allelic, and yet they differ by multiple amino acid substitutions. The possibility that the polymorphism is a regulatory one, involving polymorphic expression of duplicate genes, can be explored with nucleic acid techniques. It is also planned to test the hypothesis that change at the organismal level is more highly correlated with chromosomal rearrangement than with change at the point mutation level. These tests will be done with improved quantitative methods for comparing the karyotypes and the morphology of hundreds of vertebrate species.